准备和存储
推荐的实验流程
Western blot: Please refer to http://www.bdbiosciences.com/pharmingen/protocols/Western_Blotting.shtml
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- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
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Engagement of the T cell receptor (TcR) results in the activation of multiple intracellular src protein tyrosine kinases (PTKs) including p56 [lck] and p59 [fyn]. PTK activation induces TcRζ phosphorylation and the subsequent recruitment and activation of ZAP-70 which specifically phosphorylates SLP-76. SLP-76 interacts with multiple components of T cell signaling pathways, including Grb-2, PLC-γ, Vav, and SLAP-130 (also known as SLP-76 Associated Phosphoprotein of 130 kDa). SLAP-130 contains multiple tyrosine based motifs, proline base type I and II SH3 domain binding motifs, lysine/glutamic acid rich nuclear localization motifs, and an SH3-like domain. It associates with the SH2 domain of SLP-76 and is phosphorylated by TcR induced PTKs. This protein has also been reported to directly interact with fyn and has been referred to as fyn binding protein (FYB). Co-transfection data has shown that a limited concentration of FYB/SLAP-130 enhances TcR-induced IL-2 production, while overexpression downregulates gene expression. Thus, FYB/SLAP-130 is viewed to be a regulatory component of the fyn/SLP-76 signaling cascade in T cells.
研发参考 (4)
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Geng L, Pfister S, Kraeft SK, Rudd CE. Adaptor FYB (Fyn-binding protein) regulates integrin-mediated adhesion and mediator release: differential involvement of the FYB SH3 domain. Proc Natl Acad Sci U S A. 2001; 98(20):11527-11532. (Biology: Flow cytometry, Immunofluorescence, Immunoprecipitation, Western blot). 查看参考
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Musci MA, Hendricks-Taylor LR, Motto DG. Molecular cloning of SLAP-130, an SLP-76-associated substrate of the T cell antigen receptor-stimulated protein tyrosine kinases. J Biol Chem. 1997; 272(18):11674-11677. (Biology). 查看参考
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Oda A, Ikeda Y, Ochs HD. Rapid tyrosine phosphorylation and activation of Bruton's tyrosine/Tec kinases in platelets induced by collagen binding or CD32 cross-linking. Blood. 2000; 95(5):1663-1670. (Biology: Western blot). 查看参考
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da Silva AJ, Li Z, de Vera C, Canto E, Findell P, Rudd CE. Cloning of a novel T-cell protein FYB that binds FYN and SH2-domain-containing leukocyte protein 76 and modulates interleukin 2 production. Proc Natl Acad Sci U S A. 1997; 94(14):7493-7498. (Biology). 查看参考
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